天冬氨酰基-(天冬酰胺酰基)-β-羟化酶介导的 Notch 信号通路在小脑发育和功能中的作用。

BACKGROUND

Aspartyl-(Asparaginyl)-β-Hydroxylase (AAH) is a hydroxylating enzyme that promotes cell motility by enhancing Notch-Jagged-HES-1 signaling. Ethanol impaired cerebellar neuron migration during development is associated with reduced expression of AAH.

METHODS

To further characterize the role of AAH in relation to cerebellar development, structure, and function, we utilized an in vivo model of early postnatal (P2) intracerebro-ventricular gene delivery to silence AAH with small interfering RNA (siAAH), or over-express it with recombinant plasmid DNA (pAAH). On P20, we assessed cerebellar motor function by rotarod testing. Cerebella harvested on P21 were used to measure AAH, genes/proteins that mediate AAH's downstream signaling, i.e. Notch-1, Jagged-1, and HES-1, and immunoreactivity corresponding to neuronal and glial elements.

RESULTS

The findings demonstrated that: 1) siAAH transfection impaired motor performance and blunted cerebellar foliation, and decreased expression of neuronal and glial specific genes; 2) pAAH transfection enhanced motor performance and increased expression of neuronal and glial cytoskeletal proteins; and 3) alterations in AAH expression produced similar shifts in Notch-1, Jagged-1, and HES-1 protein or gene expression.

CONCLUSIONS

The results support our hypothesis that AAH is an important mediator of cerebellar development and function, and link AAH expression to Notch signaling pathways in the developing brain.

背景

天冬氨酰基-（天冬酰胺酰基）-β-羟化酶（AAH）是一种羟化酶，通过增强 Notch-Jagged-HES-1 信号来促进细胞迁移。发育过程中乙醇损害小脑神经元迁移与 AAH 表达降低有关。

方法

为了进一步研究 AAH 在小脑发育、结构和功能中的作用，我们利用体内模型，在出生后第 2 天（P2）进行侧脑室基因递送，用小干扰 RNA（siAAH）沉默 AAH，或用重组质粒 DNA（pAAH）过表达 AAH。在 P20，我们通过转棒试验评估小脑运动功能。在 P21 收获的小脑用于测量 AAH、介导 AAH 下游信号的基因/蛋白，即 Notch-1、Jagged-1 和 HES-1，以及对应神经元和神经胶质元素的免疫反应性。